Recent studies from our laboratory demonstrate that obese animals with the metabolic syndrome display significant coronary dysfunction that is related to activation of the renin-angiotensin system and augmented angiotensin receptor (AT1)-induced coronary microvascular vasoconstriction. Alterations in these mechanisms could significantly impair microvascular K+ channel function, in particular large conductance Ca2+-activated K+ (i.e. BKCa) channels. However, the contribution of BKCa channels to control of coronary blood flow and vascular dysfunction in the metabolic syndrome has not been examined. Therefore, the goal of this investigation is to understand the molecular and functional expression of BKCa channels which regulate coronary microvascular function and the deleterious influence of the metabolic syndrome. The proposed studies will be performed on Ossabaw swine fed a normal maintenance diet (7% kcal from fat) or an atherogenic diet (45% kcal from fat and 2% cholesterol) for 20 weeks. This high-fat/high-cholesterol diet induces many common features of the metabolic syndrome, including: obesity, insulin resistance, impaired glucose tolerance, dyslipidemia, hyperleptinemia, hypertension and neointimal hyperplasia. Using this clinically-relevant model, we propose to test the central hypothesis that impaired functional expression of BKCa channels significantly contributes to coronary microvascular dysfunction in the metabolic syndrome. To accomplish our goal we propose to examine the following three specific aims: 1) Contribution of BKCa channels to local metabolic and ischemic coronary vasodilation is diminished in Ossabaw swine with the metabolic syndrome. 2) Molecular expression of BKCa channel 1- and 21-subunits are altered and functional expression of BKCa current diminished in coronary microvascular smooth muscle cells from Ossabaw swine with the metabolic syndrome. Experiments in Aims 1 and 2 will also examine the mechanistic link between AT1/AT2 receptor signaling and physiologic regulation of coronary BKCa channels. 3) Chronic inhibition of the renin- angiotensin system improves local metabolic and ischemic coronary vasodilation as well as functional expression of BKCa channels in Ossabaw swine with the metabolic syndrome. The significance of our research is that coronary vascular dysfunction is likely an important contributor to increased cardiovascular morbidity and mortality in patients with the metabolic syndrome. The experimental design of the proposed studies is unique in that the assembled research team will perform complementary in vitro (e.g. immunocytochemistry, real-time PCR, Western blot, patch-clamp electrophysiology, isolated coronary microvessels) and in vivo (studies in conscious, chronically instrumented and anesthetized, open chest swine) experiments to integratively examine each of the proposed aims. Our results stand to offer novel insight into the mechanisms of obesity-related coronary vascular disease that could lead to novel therapies to reduce the incidence to coronary and cardiac complications in this ever increasing patient population. PUBLIC HEALTH RELEVANCE: Obesity and its related disorders have reached epidemic proportions in the United States as approximately 65% of US adults are estimated to be overweight or obese. These obese patients have significantly elevated morbidity and mortality to many cardiovascular diseases including: hypertension, stroke, coronary artery disease, cardiomyopathies, myocardial infarction, congestive heart failure, and sudden cardiac death. The goal of our research is to delineate the precise mechanisms of obesity-related cardiovascular disease and thus establish a rationale for therapeutic intervention.